Abstract

The study was conducted on 40 clinically healthy repeat breeding crossbred
cattle (15 with anovulation, 15 with delayed ovulation and 10 with normal
ovulation). The serum concentrations of calcium (Ca), inorganic phosphorus (IP),
magnesium (Mg), manganese (Mn), zinc (Zn), copper (Cu) and iron (Fe) during the
oestrus were compared among the animals with normal and impaired ovulations.
Ovulatory status of the experimental animals was determined through per rectum
examination and blood samples were collected during the oestrus.

The concentrations of Ca (mg %), IP (mg %) and Zn (ppm) varied significantly
(P<0.01) among the animals with normal ovulation (9.58±0.49, 5.45±0.15 and
1.78±0.02), anovulation (6.48±0.13, 4.45±0.14 and 0.85±0.03) and delayed
ovulation (7.36±0.13, 4.96±0.09 and 1.41±0.04). The concentrations of Mg (mg %)
and Mn (ppm) were found to be significantly (P<0.01) lower in the animals with
anovulation (2.64±0.04 and 0.14±0.01) compared to the animals with normal
(3.12±0.05 and 28±0.01) and delayed ovulation (2.98±0.06 and 0.23±0.01). In
contrast, Cu and Fe concentrations did not differ significantly among the
different ovulatory groups.

The results indicated that Ca, IP and Zn were probably crucial for the normal
ovulatory process and Mg and Mn below the optimum level might have caused
anovulation in crossbred cattle. However, Cu and Fe might have played little
role in the ovulatory process in crossbred cattle.

Introduction

Repeat breeding is one of the major problems in dairy cattle that affects
fertility and in turn incurs great economic loss to farmers. Ovulatory
disturbance is one of the major causes of repeat breeding in crossbred dairy in
India (Ibraheem Kutty and Ramchandran 2003). Mineral imbalance or deficiencies
may cause repeat breeding in cattle (Rupde et al 1993, Das et al 2002).
Variations in the level of trace elements have been found to be associated with
reproductive disorders (Jain and Madan 1984, Jain 1993). The deficiency of a
particular element may influence the level of other elements in the body fluid
and the functional characteristics of endocrine glands, especially the
hypophyseal-gonadal axis (Bhaskaran and Abdullakhan 1981).

Calcium (Ca) plays an important role in gonadotropic regulation of ovarian
steroidogenesis (Carnegie and Tsang 1984). Marginal deficiency of phosphorus
cause disturbance in the pituitary-ovarian-axis including ovulation (Bhaskaran
and Abdullakhan 1981). Zinc (Zn) deficiency may reduce GnRH secretion that
eventually leads to the arrest of ovulation (Kaswan and Bedwal 1995). Lack of
manganese (Mn) may inhibit the synthesis of cholesterol and its precursors that
in turn may limit the synthesis of sex hormones and possibly other steroids (Doisey
1973). Copper (Cu) has a significant role in maintaining optimum fertility as it
is involved in FSH, LH and estrogen activity (Desai et al 1982). A low level of
serum iron (Fe) has been found to influence normal gonadal activity possibly by
causing improper hormonal output and deterioration of general physiological
function (Sharma et al 1986).

At present, the information on blood mineral attributes during the oestrus in
repeat breeding crossbred cattle with impaired ovulation are scanty. The present
study was conducted to compare the serum level of Ca, inorganic phosphorus (IP),
magnesium (Mg), Mn, Zn, Cu and Fe during the oestrus in repeat breeding
crossbred cattle with normal and impaired ovulations.

Materials and methods

Experimental animals

The study was conducted on 40 clinically healthy repeat breeding crossbred
cattle (15 with anovulation, 15 with delayed ovulation and 10 with normal
ovulation). The experimental animals were selected from private farms located in
Khanapara, Guwahati, Assam, after confirming the status of ovulation (normal,
delayed and anovulation) through per rectum examination. The animals were
examined per rectum from the onset of oestrus at 12 h interval until 72 h to
detect ovulation. The animals were also examined per rectum on day 10 following
the onset of oestrus to determine the presence of a mature corpus luteum (CL).
Normal ovulation was considered, when corpus haemorrhagicum was detected within
36 h and a mature CL was present on day 10 following the onset of oestrus.
Anovulation was considered if corpus haemorrhagicum was absent till 72 h or a
mature CL was absent on day 10 following the onset of oestrus. Delayed ovulation
was considered, when Graafian follicle was present till 36 h and a mature CL was
present on day 10 following the onset of oestrus. The animals were free
from any anatomical abnormality of reproductive organs. The animals were
maintained under stallfed condition and fed adequate green fodders, straw,
concentrate and
ad libitum drinking water.

Sample collection and analysis

Blood samples were collected from the animals during the oestrus and serum was
separated and stored at -20oC until further analysis. The
concentrations of Ca (Webster 1962), IP (Gomori 1942) and Mg (Andreasen 1957) in
serum were estimated colorimetrically according to previously described methods.
The concentrations of Mn, Zn, Cu and Fe in serum were estimated using a Flame
Atomic Absorption Spectophotometry (Solar, AA Series Spectrometer, Thermo
Electron Corporation, UK) according to previously described methods (Fick et al
1979).

Statistical analysis

The variation in the serum concentration of different minerals was analyzed by
ANOVA using general linear model procedure (SPSS 1999). The model included the
different ovulatory groups as source of variation. The Student Newman Keuls test
was used to isolate means when effect was found significant (SPSS 1999).

The concentrations of Ca and IP were found to vary
significantly (p<0.01) among the groups. The concentrations of Ca and IP
were found to be lower in the animals with anovulation and delayed ovulation
compared to the animals with normal ovulation. Ca plays an important role in the
gonadotropic regulation of ovarian steroidogenesis (Carnegie and Tsang 1984) and
regulation of the membrane potential of oocytes. It is also suggested that Ca is
involved in the disruption of cumulus cell cohesiveness by regulating the number
of gap junctions between the cells (Peracchia 1978), which contributes to the
process of ovulation. Bhaskaran and Patil (1982) have suggested that marginal
deficiency of phosphorus could cause disturbances in the pituitary-ovarian axis
including ovulation. Moreover, a disturbed calcium-phosphorus ratio has a
blocking action on the pituitary gland and consequently on the ovarian function
(Herrick
and John
1977).
The results of this study indicate that the low Ca and IP levels might be
related to the ovulatory disturbances in crossbred cattle.

The concentration of Mg was found to be significantly (p<0.01) lower in
the animals with anovulation. Though the concentration did not differ among the
animals with delayed and normal ovulation, the concentration was found to be
lower in the animals with delayed ovulation. Although Mg does not have any
direct role in reproduction, it is involved in many of the enzymatic reactions
catalyzed by ATP associated enzymes. Moreover, it influences the absorption of
Ca and P (Sharma et al 2004). Therefore, Mg imbalance may influence reproductive
efficiency indirectly. The results of our study also revealed that the low Mg
level was associated with anovulation.

Variations in the serum concentration of Mn, Zn, Cu and Fe at oestrus in repeat
breeding crossbred cattle with normal ovulation, delayed ovulation and
anovulation are depicted in figure 2.

The concentrations of Mn and Zn were found
to vary significantly (p<0.01) among the different ovulatory groups. The
concentration of Mn was found to be significantly lower in the animals with
anovulation. Though the concentrations did not differ among the animals with
delayed and normal ovulation, these concentrations were found to be lower in the
animals with delayed ovulation. Mn has a role in steroid synthesis (Keen and
Zidenberg-cheer 1990) and it is associated with impaired reproductive functions
such as anoestrus, delayed ovulation and repeat breeding (Corah 1996). Delayed
ovulation has been induced experimentally in dairy cows by withholding Mn over a
long period (Rojas
et al
1965). The results of the present study indicate that Mg and Mn might have
played some role in the normal ovulatory process. The concentration of Zn was
found to be significantly (p<0.01) lower in the animals with anovulation
and delayed ovulation compared to the animals with normal ovulation. Zn
deficiency may lead to the reduction in GnRH secretion by hypothalamus and
eventually lead to decreased LH and FSH levels and arrest of ovulation (Kaswan
and Bedwal 1995). Apagar (1985) also has demonstrated that an optimum Zn level
is essential for maintaining the activity of LH to induce normal ovulation. The
results of the present study indicate that the low Zn level might have caused
ovulatory disturbances in crossbred cattle.

The concentrations of Cu and Fe did not differ significantly among the various
ovulatory groups. It is evident that in ruminant, Fe deficiency is rare in
grazing and well managed animals (Hidiroglou 1979, Puls 1994). Previous reports
also indicate that Cu deficiency does not influence the normal fertility in
cattle (Hunter 1977,
Whitaker 1982). The results of the present study indicate that probably Cu and
Fe did not play a crucial role in the ovulatory process of the experimental
animals.

Conclusions

The concentrations of Ca, IP and Zn
varied significantly among the different ovulatory groups. The concentrations of
Mn and Mg were found to be significantly lower in the animals with anovulation,
but did not differ in the animals with normal and delayed ovulation. In
contrast, the concentrations of Cu and Fe did not differ significantly among the
different ovulatory groups. The results indicated that in crossbred cattle, Ca,
IP and Zn were probably crucial for the normal ovulatory processes, and Mg and
Mn below the optimum level might have caused anovulation. However, Cu and Fe
might have played a little role in the ovulatory process in crossbred cattle.

References

Andreasen E 1957
On the determination of magnesium in serum and urine by the titan yellow method.
Scandinavian Journal of Clinical and Laboratory Investigation 9: 138-143

Carnegie J A and Tsang B K 1984
The calcium calmodulin system; participation in the regulation of
steroidogenesis activity at different stages of granulose cell differentiation.
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Ibraheem kutty C and Ramachandran K
2003 Bovine infertility: a
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